Bacteria communicate with one another and coordinate the behavior of the community by means of secretion and detection of small signaling molecules, and this phenomenon is termed quorum sensing. The proposed research will investigate how the information from an extracellular signaling molecule is transduced across a membrane to regulate gene expression. Specifically, the interactions between the newly discovered and widespread bacterial AI-2 autoinducer molecule and its receptor LuxPQ in Vibrio harveyi will be characterized. The specific aims are: 1. Identify mutations in LuxP that affect interaction with AI-2 and with LuxQ. Gain of function mutations will be generated in LuxP that mimic the AI-2-bound form. Dominant LuxP mutations will also be identified that are unable to interact with AI-2 or with LuxQ. 2. Identify mutations in the periplasmic domain of LuxQ that affect interaction with LuxP. A PCR-based mutagenesis strategy will be used to generate mutations that enhance LuxQ's interaction with LuxP. Mutations will also be isolated in LuxQ that suppress dominant mutant alleles of LuxP isolated in Aim 1. 3. Test AI-2 analogs for agonist or antagonist activity. The binding affinities of each synthetic analog for purified LuxP will be measured and compared to AI-2 activity in vivo. [unreadable] [unreadable]